Sensitive High-performance Liquid Chromatographic Method Research Articles

Stability Indicating HPLC Method for In-vitro Determination of Pantoprazole Sodium and its Degradation Products in Simulated Gastric and Intestinal Fluids

Background: A high-performance liquid chromatography (HPLC) method was developed for the determination of Pantoprazole Sodium (PPZ) in the presence of its degradation products. The degradation of PPZ was studied in simulated intestinal fluid (SIF) and simulated gastric fluids (SGF) in various temperature conditions. Aim: This study aimed to establish a simple, sensitive, and rapid RP HPLC method for in-vitro determination of Pantoprazole Sodium and its degradation products in simulated gastric and intestinal fluids. Objective: Pantoprazole is acid labile drug. In order to determine pantoprazole in various oral dosage forms, the stability-indicating assay of PPZ was performed in phosphate buffer (pH 6.8) representing simulated intestinal fluid (SIF) and in 0.1 molars (M) Hydrochloric acid (HCl) as simulated gastric fluid (SGF) at two different temperature conditions, i.e., 25°C and 0°C, respectively. Method: Pantoprazole sodium was obtained from the Akums laboratory in Haridwar. The analysis was performed by high-performance liquid chromatography (HPLC), Shimadzu, equipped with two LC-10 AD VP solvent-delivery modules, a SPD-10A UV–-visible detector, and a manual injector valve with 20 μL sample loop. Phenomenex ODS analytical column (150 mm × 4.6 mm i.d., 5 μm particles) was done under reversed-phase partition chromatographic conditions. The mobile phase was phosphate buffer and acetonitrile (ACN) of pH 7.4, respectively, optimized in a 70:30 (v/v) ratio followed by filtration through a 0.45 μm membrane filter and degassed by ultrasonicator before use. The mobile phase was delivered at the flow rate of 2 mL/min. The various parameters, such as linearity, accuracy and precision of the analytical method, were studied. Result: The standard curve of PPZ was linear (R2>0.99) over the concentration range of 5-30 μg/mL, and the relative standard deviation (RSD) values for intra-day and inter-day variations were in the range of 1.0-1.8%. The range of RSD was within ±2. Conclusion: The stability of PPZ in aqueous solution was pH dependent. The rate of degradation increases with decreasing pH. The pH stability of pantoprazole was studied at the above-mentioned temperature conditions. The PPZ peaks were analyzed by comparing them with fresh samples and were stable in SIF solution after 24 hours elapsed time at pH 6.8. The obtained degraded peaks in SGF (pH 1) were successfully separated from the PPZ.

Investigating a Simple and Sensitive High-performance Liquid Chromatography Method for Simultaneous Determination of Apigenin, Catalpol, and Gallic Acid Contents in Plantago Lanceolata L. and Plantago Major L.

Investigating a Simple and Sensitive High-performance Liquid Chromatography Method for Simultaneous Determination of Apigenin, Catalpol, and Gallic Acid Contents in Plantago Lanceolata L. and Plantago Major L.

Development and validation of an effective and sensitive technique for nitrate determination in fruits and vegetables using HPLC/PDA.

This study aims to develop an effective and sensitive HPLC (High Performance Liquid Chromatography) method to determine the nitrate concentration in fruits and vegetables (F & V) using a C18 column (ZORBAX Eclipse XDB-C18, 80Å, 250 × 4.6mm, 5μm (Agilent Technologies)) maintained at 40 0C, a mobile phase made up of methanol and buffer (pentane sulfonic acid sodium salt solution), and a Photo Diode Array Detector (PDA) at 225nm. The developed method is validated in terms of selectivity, linearity, accuracy, precision, suitability, the limit of detection (LOD), and the limit of quantification (LOQ) according to the European Union Decision 2002/657/EC. The result revealed that a ratio of 30: 70 of the organic modifier methanol and buffer with pH 2.8 shows the highest efficiency. The calibration curve shows linearity with a correlation coefficient (r) of 0.9985. The LOD and LOQ were found to be 2.26mg/kg and 7.46mg/kg. The recovery was in the range of 98.96-100.21%. Moreover, the greenness assessment scores of different approaches (eco-scale score of 76, AGREE score of 0.71, and few red shades in GAPI portray) were at a very excellent level. Thus, our developed method is fully validated and can determine the nitrate content in F & V.

Assessment on compatibility and safety of labels for pharmaceutical packaging

Although the secondary packing materials do not directly contact the finished drug products, compound migration may still happen between them. To ensure drug quality and safety, extractables and leachables of the packing materials should be analyzed. In this study, 2,6-di-tert-butyl-4-methylphenol (BHT) was first found in the labels for pharmaceutical packaging. For the identification of the compound, a strategy combining high performance liquid chromatography (HPLC), ultra-performance liquid chromatography-quadrupole time-of-flight mass (UPLC-Q-TOF-MS) and nuclear magnetic resonance (NMR) spectroscopy was utilized. Afterwards, a effective and sensitive HPLC method for quantification of BHT was developed and validated. Finally, a toxicological risk assessment of BHT was performed to ensure the safety of drugs.

Preparation, characterization, and pharmacokinetics of oridonin-loaded liposomes.

The aim of this study was to prepare oridonin liposomes and evaluate the physicochemical characteristics and pharmacokinetics in rats. A three-level, three-factor Box-Behnken design was used to optimize the preparation of oridonin liposomes. A highly sensitive high-performance liquid chromatographic quantification method using ultraviolet detection was established and validated for the determination of oridonin in rat plasma. Twelve Sprague-Dawley rats were randomly assigned and injected with 15 mg/kg of oridonin or oridonin liposomes via the tail vein. Pharmacokinetic parameters were estimated using a compartmental modeling approach using PKsolver software. The optimum conditions were as follows: soybean phospholipids/cholesterol ratio, 3.9:1; soybean phospholipids/drug ratio, 8.5:1; and soybean phospholipid concentration, 1.1%. Under these conditions, the mean particle size, polydispersity index, zeta potential, and encapsulation efficiency of oridonin liposomes were 170.5nm, 0.246, -30.3mV, and 76.15%, respectively. The pharmacokinetic results showed that liposomes could significantly prolong the elimination half-life (from 2.88 ± 0.55 to 13.67 ± 3.52 h), increase the area under the concentration-time curve (from 1.65 ± 0.17 to 6.22 ± 0.83 μg h/ml), and decrease the clearance (from 6.62 ± 1.38 to 1.96 ± 0.24L/kg h). The oridonin liposomes increased the elimination half-life and area under the concentration-time curve and provided a reference for the development of drugs with a short half-life.

Validation of a LC-UV method for determination of tildipirosin in goat milk

A simple, rapid, low-cost, and sensitive high-performance liquid chromatographic method was developed to determine tildipirosin in milk goat. Milk samples were precipitated with acetonitrile, and after evaporation, tildipirosin was determined by reverse-phase chromatography with an ultraviolet detector set at a wavelength of 289 nm. Tildipirosin was separated on a Zorbax Eclipse XDB-C18 column, 150 x 3.0 mm, 5 μm with gradient chromatographic elution. The retention times for tildipirosin and tylosin tartrate were 4.4 min and 10.5 min, respectively. Calibration curves were ranged from 100 to 2500 µg/L. The lower limit of detection was 75 µg/L, and the lower limit of quantitation was 100 µg/L. The accuracy and precision were always <15% except for LOQ < 20%. Mean recovery was 95.6 %. This procedure can be applied to perform pharmacokinetic studies in lactating animals and be useful to detect residues in dairy products.

Identification and quantification of extractables and leachables in laminated film and pouches for pharmaceutical packaging

Extractables and leachables from pharmaceutical packaging material can potentially be detrimental, which may affect the safety of the drug products. In this study, two compounds were found to be the possible extractables from the secondary packaging material. A strategy combining high performance liquid chromatography (HPLC), fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS) and nuclear magnetic resonance (NMR) spectroscopy was utilized for identification of the two compounds. Afterwards, a simple and sensitive HPLC method was developed and validated for the simultaneous quantification of both. The results indicated that this method was suitable for quantificating the two extractables in the pharmaceutical packaging material.

Chromatographic Analysis of Aqueous Humor of Bupivacaine in Different Administration Approaches.

Different administration approaches were investigated for the selection of bupivacaine administration type and a sensitive high-performance liquid chromatographic (HPLC) method has been developed. Developed method was validated and applied for the determination of bupivacaine in rabbit aqueous humor. The separation was achieved using a XTerra, C8 (250×8 mm i.d., particle size 5μm) analytical column with a mobile phase consisted of acetonitrile and sodium dihydrogen phosphate (pH = 3.0, 20mM; 30:70, v/v). Bupivacaine detection was performed by Diode Array detector (DAD) at 220 nm. The retention times for bupivacaine is 15.886 min. HPLC-DAD method was linear in the range of 75-4000 ng/mL. The limit of detection was 25 ng/mL and the limit of quantification of bupivacaine was found to be 75 ng/mL (relative standard deviation, RSD ≤ 15%, n = 6). In intra-day and inter-day precision and accuracy analysis, the RSD was found to be in the range of 0.96 and 7.98%, the bias values were 0.64 and 3.33%. Method was carried out for three different type of bupivacaine application because of the investigation of effective drug administration. Twenty aqueous humor samples were in the range of 0.642 and 5.124 μg/mL.

ANALYTICAL METHOD VALIDATION OF GLICLAZIDE RELATED SUBSTANCES BY HIGH PERFORMANCE LIQUID CHROMATOGRAPHY METHOD

Objective: The present study was undertaken with the objective of method validation of a rapid, simple, cost-effective HPLC method for the determination of related substances of Gliclazide. Methods: A simple, rapid, and specific method for analysis of gliclazide by a sensitive high-performance liquid chromatographic method is described. Validation of the method is carried out by USP and ICH guidelines. The method was validated for parameters like accuracy, precision, linearity, specificity, robustness, and system suitability. These proposed methods are suitable for the determination of title drugs in quality control laboratories in the pharmaceutical industries. Results: The mobile phase used for the chromatographic runs consisted of (450 ml) of acetonitrile and (550 ml) of water. The separation was achieved on the LiChroCART Supersher RP-8 column, (250 mm × 4.0 mm, 5 µm), using isocratic mode. Drug peaks were well separated and were detected by a UV detector at 235 nm, the method was linear at the concentration range. Gliclazide limit of detection (LOD) and limit of quantification (LOQ) was 0.003 and 0.01 while LOD and LOQ for Impurity-F were 0.003 and 0.01 respectively. Conclusion: The presented validated method is rapid, economic, simple, accurate, sensitive, robust, specific, and linear. It can be used for routine analysis of gliclazide.

Development and Validation of an Improved HPLC-UV Method for the Determination of Tildipirosin in Horse Plasma

Abstract A simple, rapid, low-cost, and sensitive high-performance liquid chromatographic method was developed to determine tildipirosin in horse plasma. Plasma samples were extracted with diethyl ether, and after evaporation, tildipirosin was determined by reverse-phase chromatography with an ultraviolet detector set at a wavelength of 289 nm. Tildipirosin was separated on a Zorbax Eclipse XDB-C18 column, 150 x 3.0 mm, 5 μm with gradient chromatographic elution. The retention times were 3.0 min and 6.4 min for tildipirosin and tylosin tartrate, respectively. The total run time was 9 minutes in this method. Calibration curves ranged from 0.1 to 3 μg/mL. The lower limit of detection for plasma was0.035μg/mL, and the lower limit of quantitation was 0.1 μg/mL. Both accuracy and precision were always < 12% exce pt for LLOQ < 20%. Mean recovery was 99.5 %. This procedure can be applied to determine tildipirosin concentrations in plasma and be useful to perform pharmacokinetic studies.

Development and validation of UV chromatographic method for quantification of copper and copper nanoparticles in different matrices and pharmaceutical products

Background The applications of Cu and CuNPs based on the earth-abundant and inexpensive Cu metal have generated a great deal of interest in recent years, including medical applications. A novel, specific, precise, accurate and sensitive reverse-phase high-performance liquid chromatography (RP-HPLC) method with UV detection has been developed and validated to quantify copper (Cu) and copper nanoparticles (CuNPs) in different biological matrices and pharmaceutical products. Methods The developed method has been validated for linearity, precision, sensitivity, specificity and accuracy. Cu concentration was detected in pharmaceutical products without an extraction process. Moreover, liver, serum and muscle tissues were used as biological matrices. High Cu recovery in biological samples was afforded by using citric acid as a green chelating agent, exact extraction time and pH adjustment. Cu pharmaceutical and biological samples were eluted by acetonitrile: ammonium acetate (50 mM) with 0.5 mg/ml EDTA (30:70 v:v) as an isocratic mobile phase. EDTA reacted with Cu ions forming a Cu-EDTA coloured complex, separated through the C18 column and detected by UV at 310 nm. Results The developed method was specific with a short retention time of 4.95 min. It achieved high recovery from 100.3% to 109.9% in pharmaceutical samples and 96.8–105.7% in biological samples. The precision RSD percentage was less than two. The method was sensitive by achieving low detection limits (DL) and quantification limits (QL). Conclusion The validated method was efficient and economical for detecting Cu and CuNPs by readily available chemicals as EDTA and Citric acid with C18 column, which present the best results on RP-HPLC.

Development and Validation of a Novel HPLC Method to Analyse Metabolic Reaction Products Catalysed by the CYP3A2 Isoform: In Vitro Inhibition of CYP3A2 Enzyme Activity by Aspirin (Drugs Often Used Together in COVID-19 Treatment).

Aspirin (also known as acetylsalicylic acid) is a drug intended to treat fever, pain, or inflammation. Treatment of moderate to severe cases of COVID-19 using aspirin along with dexamethasone has gained major attention globally in recent times. Thus, the purpose of this study was to use High-Performance Liquid Chromatography (HPLC) to evaluate the in vitro inhibition of CYP3A2 enzyme activity using aspirin in rat liver microsomes (RLMs). In this study, an efficient and sensitive HPLC method was developed using a reversed phase C18 column (X Bridge 4.6 mm × 150 mm, 3.5 µm) at 243 nm using acetonitrile and water (70:30 v/v). The linearity (r2 > 0.999), precision (<15%), accuracy and recovery (80–120%), limit of detection (5.60 µM and 0.06 µM), limit of quantification (16.98 µM and 0.19 µM), and stability of the newly developed method were validated for dexamethasone and 6β-hydroxydexamethasone, respectively, following International Conference on Harmonization (ICH) guidelines. This method was applied in vitro to measure CYP3A2 activity. The results showed that aspirin competitively inhibits 6β-hydroxylation (CYP3A2 activity) with an inhibition constant (Ki) = 95.46 µM and the concentration of the inhibitor causing 50% inhibition of original enzyme activity (IC50) = 190.92 µM. This indicated that there is a minimal risk of toxicity when dexamethasone and aspirin are co-administrated and a very low risk of toxicity and drug interaction with drugs that are a substrate for CYP3A2 in healthcare settings.

High-performance liquid chromatography quantitative analysis of ephedrine alkaloids in Ephedrae Herba on a perfluorooctyl stationary phase.

Ephedrae Herba is one of the most commonly used herbal medicines, and it has been shown that most of the clinical efficacy for cold and asthma is exerted by its alkaloidal components. A simple and sensitive high-performance liquid chromatography method was developed using a perfluorooctyl column for the simultaneous determination of five alkaloids (norephedrine, norpseudoephedrine, ephedrine, pseudoephedrine, and methylephedrine) in Ephedrae Herba. The mobile phase comprising acetonitrile and 15mM ammonium trifluoroacetate was used to elute the targets in isocratic elution mode. The method was validated for linearity (R2 >0.999), repeatability, intraday and interday precision, recoveries with trueness (93.87-110.99%), limits of detection (5.35-5.76µg/mL), and limits of quantification (20µg/mL). The quantitative results revealed that the developed method was precise and accurate. Then it was successfully applied to determine the difference in the contents ofthree batches of Ephedrae Herba from three pharmaceutical companies.

Analysis of cycloheximide in rat specimens using liquid chromatography with tandem mass spectrometry detection

The development of a selective and sensitive high-performance liquid chromatographic tandem mass spectrometric method for the determination of cycloheximide (CHX) in rat blood and plasma is described. The extraction of CHX and colchicine as internal standard from blood fluid (0.1 mL) was achieved using n-hexane: dichloromethane: isopropanol (20:10:1 v/v/v). The mobile phase, a combination of methanol:10 mM ammonium acetate (85:15, v/v), was pumped at 0.2 mL/min through a C18 analytical column with a run time of 3.5 min. Detection was carried out by electrospray positive ionization mass spectrometry in the multiple-reaction monitoring (MRM) mode. The assay exhibited excellent linearity (r2 > 0.999) in peak area response over the concentration ranges of 2–1000 ng CHX /mL blood fluid. The mean absolute recoveries for 20, 100 and 500 ng/mL CHX in blood fluid using the present extraction procedure were > 97%. The intra- and inter-day coefficients of variation in the plasma and blood and mean error were < 13% at different concentrations. Samples had limited stability at room temperature, and speedy processing is needed. After intravenous administration, rats had measurable concentrations of CHX for up to 24 h after dosing with 1 mg/kg of cycloheximide. The method displayed a high caliber of sensitivity and selectivity for detecting very low concentrations of CHX in rats.

Comparative Pharmacokinetic Studies of Marketed and Microsponges Gel Loaded with Diclofenac Diethylamine in Rabbits

Oral administration of the non-steroidal anti-inflammatory drug, Diclofenac Diethylamine (DDEA) is often associated with gastrointestinal ulcers, bleeding and extensive first-pass hepatic metabolism. As an alternative to oral administration, formulated microsponges-based gel of DDEA was developed for topical administration, to quantify diclofenac diethylamine in plasma of rabbits for this a sensitive Reverse Phase High-Performance Liquid Chromatography (RP-HPLC) method was developed using carbamazepine as Internal standard (IS) and DDEA (pure drug) was provoked on Hypersil RP C18 column (250mm × 4.6mm 5µm) using a mobile phase mixture of potassium dihydrogen buffer pH 2.5 and acetonitrile in the ratio of 30:70 v/v at an isocratic flow rate of 1mL/min. The drug peak area was detection and found at 276nm. The retention time of DDEA was found to be 5.3 min. The calibration curve was linear over the concentration range of 50-750ng/ml of DDEA. This method was accurate for quantitative estimation of the drug from the marketed gel and optimized microsponge gel. The main of investigation is to compare pharmacokinetic profile of diclofenac diethylamine in pharmaceutical dosage forms (marketed gel and microsponges gel) using WinNonlin software version 8.1.

A novel liquid chromatography-fluorescence method for the determination of delafloxacin in human plasma.

Delafloxacin is a novel fluoroquinolone antibiotic that was approved by the European Medicine Agency to treat bacterial infections of the skin and underlying tissues, and community-acquired pneumonia. Despite being in the market since 2019 in the European Union, there is no published liquid chromatography-fluorescence method for delafloxacin quantification in biological samples. A novel, rapid, and sensitive high-performance liquid chromatographic method was developed to determine delafloxacin in human plasma using its native fluorescence. Plasma delafloxacin concentrations were determined by reverse-phase chromatography with fluorescence detection at 405/450nm of excitation/emission wavelengths. Delafloxacin was separated on a Kromasil C18 column 250 × 4.6mm id, 5 µm using isocratic elution. The mobile phase was a mixture of 0.05% trifluoroacetic acid/acetonitrile (52/48). Retention times were 5.4 and 11.6 min for delafloxacin and valsartan (internal standard), respectively. Regression calibration curves were linear over the range of 0.1-2.5 µg/mL. The lower limit of detection was 0.05 µg/mL, and the lower limit of quantification was 0.1 µg/mL. Accuracy and precision were always<11%, and the limit of quantification was<16%. Mean recovery was 98.3%. This method can be applied to determine delafloxacin in human plasma and could be useful to perform pharmacokinetic studies.

Effect of Sijunzi Pills on Pharmacokinetics of Omeprazole in Beagle Dogs by HPLC-UV: A Herb-Drug Interaction Study.

A sensitive high-performance liquid chromatography (HPLC-UV) method for determination of omeprazole in beagle dog plasma was developed and to investigate the effect of Sijunzi pills (SJZPs) on the pharmacokinetics of omeprazole in beagle dogs. The beagle dog plasma was extracted with ethyl acetate and n-hexane under alkaline conditions. Omeprazole and internal standard (IS, fluconazole) were separated on an XDB-C18 column, and acetonitrile and 0.1% trifluoroacetic acid were used as the mobile phase. Omeprazole and IS were detected by using a diode array detector. This experiment adopts the experimental design of double-cycle self-control. In the first cycle (group A), six beagle dogs were given omeprazole 0.67 mg/kg orally in a single dose. In the second period (group B), the same six beagle dogs were orally given SJZPs 0.2 g/kg twice a day for 7 consecutive days, and then, omeprazole was orally given. At the different time points after omeprazole was given in the two periods, the blood samples were collected. The concentration of omeprazole was detected by the developed HPLC method. DAS 2.0 was used to calculate the pharmacokinetic parameters of omeprazole. Under the current experimental conditions, this UPLC method showed good linearity in the detection of omeprazole. Interday and intraday precision did not exceed 10%, and the range of accuracy values were from −1.43% to 2.76%. The results of extraction recovery and stability met the requirements of FDA approval guidelines of bioanalytical method validation. The Cmax of omeprazole in group B was 61.55% higher than that in group A, and the AUC(0−t) and AUC(0−∞) of omeprazole in group B were 63.96% and 63.65% higher those that in group A, respectively. At the same time, the clearance (CL) and apparent volume of distribution (Vd) decreased in group B. In this study, an HPLC method for the determination of plasma omeprazole concentration was established. SJZPs could inhibit the metabolism of omeprazole and increase the concentration of omeprazole in beagle dogs. It is suggested that when SJZPs are combined with omeprazole, attention should be paid to the herb-drug interactions and possible adverse reactions.

Application of Solid Phase Extraction and High-Performance Liquid Chromatography with Fluorescence Detection to Analyze Bisphenol A Bis (2,3-Dihydroxypropyl) Ether (BADGE 2H2O), Bisphenol F (BPF), and Bisphenol E (BPE) in Human Urine Samples.

In this study, we propose a simple, cost-effective, and sensitive high-performance liquid chromatography method with fluorescence detection (HPLC-FLD) for the simultaneous determination of the three bisphenols (BPs): bisphenol A bis (2,3-dihydroxypropyl) ether (BADGE 2H2O), bisphenol F (BPF), and bisphenol E (BPE) in human urine samples. The dispersive solid phase extraction (d-SPE) coupled with solid phase extraction (SPE) procedure performed well for the analytes with recoveries in the range of 74.3–86.5% and relative standard deviations (RSD%) less than 10%. The limits of quantification (LOQs) for all investigated analytes were in the range of 11.42–22.35 ng mL−1. The method was validated at three concentration levels (1 × LOQ, 1.5 × LOQ, and 3 LOQ). During the bisphenols HPLC-FLD analysis, from 6 min a reinforcement (10 or 12) was used, therefore analytes might be identified in the small volume human urine samples. The results demonstrated clearly that the approach developed provides reliable, simple, and rapid quantification and identification of three bisphenols in a urine matrix and could be used for monitoring these analytes.

Evaluation of intraocular penetration of levofloxacin by high performance liquid chromatography

AIM: To evaluate the levofloxacin eye drop into human eye penetration, levofloxacin eye drop concentrations in human ocular aqueous of 33 patients undergoing cataract surgery were measured by high performance liquid chromatography (HPLC). METHODS: Totally 33 volunteer patients who scheduled for phacoemulsification surgery received one drop of levofloxacin every 6h for 3d before and on the day of surgery, administration of drug was stopped 1h before surgery. Levofloxacin concentration in aqueous humor was measured by HPLC method with fluorescence detector. RESULTS: A simple, effective and sensitive HPLC method for determination of levofloxacin in human ocular aqueous was validated. Linearity was shown for levofloxacin concentration over a wide range of 1.95×10-3-1.50 µg/mL. The mean aqueous level of levofloxacin was 0.3399±0.03405 µg/mL. CONCLUSION: Results from the present study demonstrate that topical administration of levofloxacin 0.5% before cataract surgery with routine dose (one drop every 6h) unable to reach MIC90 for most common microorganism causing acute bacterial endophthalmitis.

Fruits of Vitex doniana sweet: toxicity profile, anti-inflammatory and antioxidant activities, and quantification of one of its bioactive constituents oleanolic acid

BackgroundVitex doniana Sweet fruit, an under-utilised crop specie of Ghana, has not been validated for its ethnomedical use in managing inflammatory conditions. Therefore, the study sought to investigate its anti-inflammatory and antioxidant activities as well as isolate and quantify one of its active constituents. Materials and methodsIn-vivo anti-inflammatory activity of the methanol fruit extract was evaluated using the carrageenan-induced oedema model in chicks. The in-vitro antioxidant property was also investigated using the 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay. The acute and subacute toxicity studies of the fruit extract were evaluated in rodent models. ResultsNo signs of autonomic and central nervous system stimulation/depression were recorded. The LD50 by oral route, was estimated to be beyond 3000 mg/kg. Subacute studies revealed an increase in red blood cell and lymphocyte counts. Liver enzymes, serum proteins and bilirubin levels did not significantly increase. The crude extracts at doses of 10, 30 and 100 mg/kg inhibited paw oedema considerably. The ethyl acetate fraction showed the highest antioxidant activity (IC50 = 99.35 ± 0.77 μg/mL). Oleanolic acid, isolated from the ethyl acetate extract, showed significant anti-inflammatory and antioxidant activities. A sensitive high-performance liquid chromatography method for the detection and estimation of oleanolic acid, as a biomarker compound for V. doniana fruit, was developed and validated for quality assurance purposes. ConclusionThe extract of V. doniana fruits possesses considerable anti-inflammatory and antioxidant properties and was non-toxic under laboratory conditions.